We focus on developing an understanding of toxic metal action in the human testis. A male factor is present about 60% of infertile couples, but underlying molecular mechanisms are largely uncharacterized. Exciting results from our current work hints at one mechanism. Lead levels were elevated markedly in testes and seminal plasma (in 25% of males in four independent populations). High lead correlated with expression of particular potassium and calcium ion channel isofomis, with poor sperm-fertilization-potential biomarkers and low fertility by IVF, artificial insemination and coitus. A significant fraction of subjects studied longitudinally switched from high lead states to low lead states, with simultaneous conversion of biomarkers from infertile to fertile and switch in potassium channel isoform expression. This suggested lead epigerietically modified testicular gene expression (at the levels of transcription and mRNA splicing) and that potassium channel isoforms could be developed as biomarkers for lead exposure. A preliminary DNA microarray study of a lead-treated "lead-resistant" rat strain identified many lead-affected genes as being involved in calcium-mediated induction of apoptosis, including a potassium channel. Supported by current somatic cell apoptosis mechanisms, this prompted our hypothesis that lead exposures produce male infertility by altering calcium homeostatsis, and a related detailed mechanism of lead action. These will be tested in a lead-treated lead-sensitive" rat strain and in humans. We will use microarrays to probe in rats for affected testicular genes with CAMP response elements and other genes involved in calcium/calmodulin-dependent protein kinase IV signaling. Controls include metal testing by atomic absorption, TUNEL estimates of apoptosis, cell type levels by histology and by cell-type-specific mRNA levels, and protein expression by Westerns. Comparison with the "lead-resistant" strain should identify lead-sensitivity" genes. We will probe for the same genes in a human clinical population, with similar controls. We will also probe for genes co-regulated with the potassium channel above. Results will test several specific steps in our proposed mechanism: verifying, negating or modifying it. Because microarrays cannot detect differential calcium channel splicing events correlated with lead effects upon human testes, this gene and other calcium transporters will be studied by immunocytochemistry, RT in situ PCR and real-time PCR. Outcome is test of hypothesis, and possible mechanism explaining infertility associated with low sperm counts or idiopathic male infertility, tools for diagnosis, and hope for treatment.